Antonie van Leeuwenhoek was the first to observe microorganisms under a microscope in 1676. Microorganisms proved harder to grow than to visualize until 1860 when Louis Pasteur first used a culture medium to grow bacteria in a laboratory.1 In the years that followed the development of antibiotics and other inhibitors, more refined and consistent nutrient sources, growth factors, and chromogenic and fluorogenic substrates allowed for the production of microbe-specific and high-quality culture media. These developments revolutionized the field of microbiology and have made the use of culture media a reliable and cost-effective tool for microbiological analyses.
Effective media formulations require water and sources of nutrients such as carbon, nitrogen, inorganic compounds, metals, and vitamins. Media formulations must be used as described in Standard Methods because even minor changes can greatly alter the recovery of the target organisms. Inaccurate measurements and inadequate mixing of media can result in media that is too concentrated or dilute, which stresses the microorganism being recovered, resulting in inaccurate results. Commercially prepared media, which require only weighing and dissolving of the powder in reagent water in the laboratory, are likely to be of consistent quality because of extensive manufacturing QA/QC requirements. Use commercially dehydrated media, when available, for consistent results.
There are several different classifications of media, determined by the media constituents. General growth media do not contain inhibitors and are designed to support the growth of a wide variety of organisms. Selective media contain constituents that promote the growth of specific microorganisms and contain inhibitors to prevent the growth of nontarget organisms. Selectivity can be obtained in many ways and to varying degrees by compounds such as antibiotics, dyes, and other inhibitory chemicals that are incorporated into the medium. Agents used to impart specificity in a medium can cause additional stress and reduced recovery, even in healthy populations of the target organisms, especially if at higher concentrations. Enrichment media are broth media designed to support and promote the growth of a specific group of organisms. Enrichment media contain inhibitors, but generally in sufficiently low amounts to avoid further stress to the target cells. Environmental organisms, already experiencing stress from the various environmental conditions and exposures, may have increased sensitivity to secondary stressors such as inhibitory compounds, and the effect is reduced detection. For example, removing rosolic acid from mFC medium was shown to result in an increased growth of fecal coliforms.2 Care must be taken when considering the most appropriate medium to be used for environmental analyses. Generally, use the least inhibitory medium possible to obtain the most accurate data.
Differential media contain components that allow for the detection and discrimination of different types of organisms. In a medium that is both selective and differential, often the medium can be used for presumptive identification of a specific group of microorganisms. Chromogenic and fluorogenic media are examples of media that are both selective and differential.
Media are available commercially in both dehydrated and prepared forms. The dehydrated media are prepared and sterilized by the laboratory in-house. A commercially prepared medium is ready to use and does not require any additional preparation. Appropriate quality control must be performed on all media (whether prepared in house or purchased commercially), and media must be stored appropriately and used by its expiration date. For media prepared onsite, review manufacturer’s instructions, product safety data sheets (SDSs) and analytical method requirements before use.
The formulations of dehydrated or prepared media are included so the user understands how the components work to detect the desired environmental organisms. Detection and isolation of environmental microorganisms is challenging because of the multiple varying parameters of a water matrix (including soils and biofilm) coupled with the unique strains of organisms present in that matrix at any given time. No single medium can be the best for all water sources and microorganisms at all times. Therefore, laboratories are encouraged to learn about the media and techniques available, and then test the methodology they intend to use across several seasons. Compare the results with a reference method to determine the most effective medium for the water matrix being examined.